Probiotics are now garnering attention as a dietary supplement owing to its medicinal and therapeutic benefits. It has shown exponential growth in the last decade and is now available in different formulations, and as probiotic enriched food and beverages. Viability is considered as an important aspect for the function of probiotics. A shelf stable, viable probiotic would always have a commercial edge over the others available in the market. Also, accurate enumeration of viable count of probiotic is important for its therapeutic activity. Probiotics are known to exist in a viable but non culturable state wherein, the standard plate count method of enumeration will not able to enumerate the spore count accurately. Several alternate methods can be used to determine the viable bacterial count which includes fluorescent in situ hybridization (FISH), polymerase chain reactions and microplate fluorochrome assay. (Enumeration of probiotic strains: Review of culture-dependent and alternative techniques to quantify viable bacteria, Catherine Davis, Journal of Microbiological Methods, Volume 103, August 2014, Pages 9-17).
The plate count method does not support precise, reproducible estimations of cell densities of probiotic strains, especially in mixed cultures (Suitability of MRS-bile agar for the selective enumeration of mixed probiotic bacteria in presence of mesophilic lactic acid cultures and yoghurt bacteria, S. Sohrabvandi, A.-M. Mortazavian, M.-R. Dolatkhnejad, A. B. Monfared, Iranian Journal of Biotechnology., 10 (2014 pp, 16-20. It estimates only the subset of viable organisms that replicate under the conditions of culture.
Hence it is necessary to develop a new method to determine the actual bacterial count of probiotic formulation. The present invention solves the above mentioned problems by disclosing a stable probiotic composition and its method of detection.
It is principle objective of the present invention to disclose a stable probiotic composition containing Bacillus coagulants MTCC 5856 exhibiting increased viability and stability over wide range of pH compared to other commercially available probiotics.
It is yet another objective of the present invention to disclose a simple, accurate, commercially viable flow cytometric method to detect and enumerate live spores and vegetative cells.
The present invention fulfils the aforesaid objectives and provides further related advantages.